In laser microdissection, in particular of biological or medical samples such as tissue biopsy samples, a sample having a thickness of generally 5 to 10 micrometers is placed upon a slide of a laser dissection microscope, whereafter dissected specimens are cut from the sample by means of a finely focused laser beam of an infrared (IR) laser diode (for example AlGaAs laser diode). To that end, a transparent thermoplastic transfer film selectively absorbing in the infrared emission spectrum of the laser diode is placed over the sample. The film is melted at well defined locations and adheres to the desired part of the sample to be cut (dissected specimen). The dissected specimen may comprise single cells or a group of cells, for example, which need to be extracted from the probe for further analysis. In other words, the dissected specimens, the size of which is of the order of about 1 to 10 micrometers, need to be collected after cutting. Here, several problems arise, in particular if a plurality of dissected specimens has to be cut and collected. The sample is, moreover, thermally affected since the IR laser beam employed for cutting transfers heat onto the sample and may, thereby, damage the sample such as, in this example, the cells.
Several methods are known in the art for collecting dissected specimens. German Patent No. DE 20100866 U describes, for example, a carrier device which freely supports the transfer film with the sample within the frame and from which the film with the dissected specimen falls down into a receptacle after having been cut by the laser beam. This method has, however, the following disadvantages. On the one hand there is the lack of referenceability of the dissected specimens because all dissected specimens fall into the receptacle in an uncontrolled manner and thus cannot be related to their original positions. Further, due to their merely gravity-induced free fall, small dissected specimens, in particular, may be lost for analysis if they get caught on parts of the microscope or the walls of the receptacle their electrostatic interactions.
U.S. Pat. No. 5,998,129 describes a laser microdissection method, wherein a desired region, for instance cellular organelles or single cells, is cut out from the surrounding tissue of a tissue sample placed upon a planar slide by means of a laser beam. The isolated cell, which is still present on the slide, is catapulted along the laser beam by an additional laser pulse and caught in a reaction receptacle. Due to the transfer of momentum from the laser beam the dissected specimens again fly into the reaction receptacle in an uncontrolled manner and can no longer be referenced. Additionally, large dissected specimens cannot be catapulted and have to be separated beforehand.
A further known method comprises the use of an adherent collection device, a so-called cap. First, several dissected specimens are cut from the sample, then the cap is lowered onto the sample and the dissected specimens are adhered to the cap. The cap with the adhered dissected specimens is raised within the microscope and removed. Although in this method the cap can take up several dissected specimens and is thus capable of working faster, the problem arises that due to the different sizes and shapes of the dissected specimens, the limited reception area and adherability of the cap and the possible inclination of the cap upon adhering the cap surface is used only insufficiently leading to an inevitable waste of the expensive caps. If, on the other hand, several dissected specimens are stacked upon another, the danger arises that the dissected specimens may fall down in an uncontrolled way and be lost. Thus, the number of samples on the cap is severely limited, which is a disadvantage since in some applications the number of dissected specimens may then not be sufficient for analysis.
It is therefore an object of the present invention to provide a method and an apparatus solving the above-mentioned problems and allowing a contamination-free, secure, efficient and cost effective collection of dissected specimens of different sizes without damaging the samples or presenting a complicated design or structure.